NMDA antagonism method

ABSTRACT

The present invention is directed to a class of 4,6-disubstituted tryptophan derivatives, 4,6-disubstituted kynurenines, their use as NMDA antagonists and to pharmaceutical compositions containing these compounds.

This is a divisional, of application Ser. No. 08/165,144, filed Dec. 10,1993, now U.S. Pat. No. 5,484,814, which is a divisional of applicationSer. No. 08/057,195 filed May 3, 1993, now abandoned, which was acontinuation of Ser. No. 07/827,188, filed Feb. 3, 1992, now abandoned,which was a continuation in part of Ser. No. 07/662,670, filed Feb. 28,1991, now abandoned, herein incorporated by reference.

The present invention is directed to a new class of excitatory aminoacid antagonists, their use in the treatment of disease states such asepilepsy, anxiety, stroke, and to pharmaceutical or diagnosticcompositions containing these excitatory amino acid antagonists. Afurther aspect of this invention is directed to the discovery of a newuse for a group of known 6-halo-tryptophan and 4-halo-kyureninederivatives.

In accordance with the present invention, it has been discovered thatthe following class of tryptophan derivatives are excitatory amino acidantagonists: ##STR1## in which X and Y are each independently selectedfrom the group consisting of Cl, Br, F, CH₃, and CH₂ CH₃ ; or apharmaceutically acceptable salt thereof.

It has also been discovered that the following kynurenine derivativesare excitatory amino acid antagonists: ##STR2## in which X and Y areeach independently selected from the group consisting of Cl, Br, F, CH₃,and CH₂ CH₃ ; or a pharmaceutically acceptable salt thereof.

Additionally, it has been discovered that the following known compoundsare excitatory amino acid antagonists: ##STR3## in which Hal isrepresented by a halogen atom, or a pharmaceutically acceptable saltthereof.

As used in this application:

a) the term "halogen" refers to a fluorine, chlorine, or bromine atom;

b) the term "pharmaceutically acceptable addition salts" refers toeither an acid addition or a basic addition salt.

The compounds of Formula Ia, Ib, IIa, and IIb can exist as eitherpharmaceutically acceptable acid addition salts or as pharmaceuticallyacceptable basic addition salts. These compounds may also exist aszwitterions.

The expression "pharmaceutically acceptable acid addition salts" isintended to apply to any non-toxic organic or inorganic acid additionsalt of the base compounds represented by Formula Ia, Ib, IIa, IIb orany of its intermediates. Illustrative inorganic acids which formsuitable salts include hydrochloric, hydrobromic, sulphuric, andphosphoric acid and acid metal salts such as sodium monohydrogenorthophosphate, and potassium hydrogen sulfate. Illustrative organicacids which form suitable salts include the mono-, di-, andtricarboxylic acids. Illustrative of such acids are for example, acetic,glycolic, lactic, pyruvic, malonic, succinic, glutaric, fumaric, malic,tartaric, citric, ascorbic, maleic, hydroxymaleic, benzoic,hydroxy-benzoic, phenylacetic, cinnamic, salicyclic, 2-phenoxy-benzoic,p-totuenesulfonic acid, and sulfonic acids such as methane sulfonic acidand 2-hydroxyethane sulfonic acid. Such salts can exist in either ahydrated or substantially anhydrous form. In general, the acid additionsalts of these compounds are soluble in water and various hydrophilicorganic solvents, and which in comparison to their free base forms,generally demonstrate higher melting points.

The expression "pharmaceutically acceptable basic addition salts" isintended to apply to any non-toxic organic or inorganic basic additionsalts of the compounds represented by Formula Ia, Ib, IIa, IIb or any ofits intermediates. Illustrative bases which form suitable salts includealkali metal or alkaline-earth metal hydroxides such as sodium,potassium, calcium, magnesium, or barium hydroxides; ammonia, andaliphatic, alicyclic, or aromatic organic amines such as methylamine,dimethylamine, trimethylamine, and picoline. Either the mono- ordi-basic salts can be formed with those compounds.

All of the compounds of Formula Ia, Ib, IIa, and IIb contain a chiralcenter and thus can exist as optical isomers. Any reference to thesecompounds or their intermediates should be construed as referring toeither a racemic mixture or an individual optical isomer. The specificoptical isomers can be separated and recovered by techniques known inthe art such as chromatography on chiral stationary phases or resolutionvia chiral salt formation and subsequent separation by selectivecrystallization. Alternatively utilization of a specific optical isomeras the starting material will produce the corresponding isomer as thefinal product.

The compounds of Formula Ia are substituted at the 4 and 6 positions asis indicated by the X and Y substituents. X and Y can be represented bythe same substituent or by differing substituents. The compounds ofFormula Ib are substituted at positions 4 and 6 as is indicated by the Xand Y substituents. X and Y can be represented by the same substituentsor differing substituents.

Examples of compounds encompassed by Formula Ia include:

4-Bromo-6-fluorotryptophan;

4-Bromo-6-chlorotryptophan;

4-Ethyl-6-bromotryptophan;

4,6-Dibromotryptophan;

4,6-Dichlorotryptophan;

Examples of compounds encompassed by Formula Ib include:

4,6-Dichlorokynurenine;

4-fluoro-6-Bromo-kynurenine;

4-chloro-6-bromokynurenine;

4,6-Dibromo-kynurenine;

6-Ethyl-4-bromokynurenine;

Examples of compounds encompassed by Formula IIa include:

6-chloro-tryptophan;

6-fluoro-tryptophan.

Examples of compounds encompassed by Formula IIb include:

4-chloro-kynurenine;

4-fluoro-kyurenine.

It is preferred for X and Y to each be represented by a halogen atom ineither the tryptophans of Ia or the Kynurenines of Ib.

The compounds of Formula Ia may be prepared using techniques andprocedures well known and appreciated by one of ordinary skill in theart. A general synthetic procedure for preparing these compounds is setforth in Scheme A. In Scheme A, all substituents unless otherwiseindicated are as previously defined. ##STR4##

Scheme A provides a general synthetic scheme for preparing compounds ofFormula Ia.

In step a, the appropriate 4,6-substituted indole-2-carboxylic acid ofstructure (1) is decarboxylated to give the corresponding4,6-substituted indole of structure (2).

For example, the appropriate 4,6-substituted indole-2-carboxylic acid ofstructure (1) is contacted with a catalytic amount of copper. Thereactants are typically contacted in a suitable organic solvent such asquinoline. The reactants are typically stirred together for a period oftime ranging from 1-5 hours and at a temperature range of from 200°-220°C. The 4,6-substituted indole of structure (2) is recovered from thereaction zone by extractive methods as is known in the art. It may bepurified by silica gel chromatography.

In step b, the appropriate 4,6-substituted indole of structure (2) isalkylated with ethyl 3-bromo-2-hydroxyiminopropanoate (3) to give theethyl 2-(hydroxyimino)-3-(4,6-substituted-3-indolyl)propanoate ofstructure (4).

For example, the appropriate 4,6-substituted indole of structure (2) iscontacted with a molar equivalent of ethyl3-bromo-2-hydroxyiminopropanoate (3) and a molar excess of a suitablebase such as potassium carbonate. The reactants are typically contactedin a suitable organic solvent such as methylene chloride. The reactantsare typically stirred at room temperature together for a period of timeranging from 10-50 hours. The ethyl2-(hydroxyimino)-3-(4,6-substituted-3-indolyl)propanoate of structure(4) is recovered from the reaction zone by extractive methods as isknown in the art. It may be purified by silica gel chromatography.

In step c, hydroxyimino functionality of the appropriate ethyl2-(hydroxyimino)-3-(4,6-substituted-3-indolyl)propanoate of structure(4) is reduced to give the 4,6-substituted tryptophan ethyl ester ofstructure (5).

For example, the appropriate ethyl2-(hydroxyimino)-3-(4,6-substituted-3-indolyl)propanoate of structure(4) is contacted with a molar excess of zinc. The reactants aretypically contacted in a suitable acidic solvent, such as acetic acid.The reactants are typically stirred together at room temperature for aperiod of time ranging from 10-100 hours. The 4,6-substituted tryptophanethyl ester of structure (5) is recovered from the reaction zone byextractive methods as is known in the art. It may be purified by silicagel chromatography.

In step d, the ethyl ester functionality of the appropriate4,6-substituted tryptophan ethyl ester of structure (5) is removed togive the corresponding 4,6-substituted tryptophan of structure (6).

In order to facilitate the purification of the tryptophan of structure(6), the amino functionality of the appropriate 4,6-substitutedtryptophan ethyl ester of structure (5) is first protected as itscarbobenzyloxy derivative.

For example, the appropriate 4,6-substituted tryptophan ethyl ester ofstructure (5) is contacted with molar equivalent of benzyl chloroformateand a slight molar excess of triethylamine. The reactants are typicallycontacted in a suitable organic solvent such as methylene chloride. Thereactants are typically stirred together at room temperature for aperiod of time ranging from 5-24 hours. The intermediate 4,6-substitutedtryptophan-carbobenzyloxy ethyl ester is recovered from the reactionzone by extractive methods as is known in the art. It may be purified bysilica gel chromatography.

The ethyl ester functionality of the appropriate intermediate4,6-substituted tryptophan-carbobenzyloxy ethyl ester is then removed togive the intermediate 4,6-substituted tryptophan-carbobenzyloxy.

For example, the appropriate intermediate 4,6-substitutedtryptophan-carbobenzyloxy ethyl ester is contacted with a molar excessof a suitable base such as lithium hydroxide. The reactants aretypically contacted in a suitable solvent mixture such astetrahydrofuran/water. The reactants are typically stirred together atroom temperature for a period of time ranging from 1-5 hours. Theintermediate 4,6-substituted tryptophan-carbobenzyloxy is recovered fromthe reaction zone by extractive methods as is known in the art. It maybe purified by silica gel chromatography.

The carbobenzyloxy functionality of the appropriate intermediate4,6-substituted tryptophan-carbobenzyloxy-carbonyl is then removed togive the 4,6-substituted tryptophan of structure (6).

For example, the appropriate intermediate 4,6-substitutedtryptophan-carbobenzyloxy is contacted with a molar excess oftrimethylsilyl iodide. The reactants are typically contacted in asuitable organic solvent such as chloroform. The reactants are typicallystirred together at room temperature for a period of time ranging from1-5 hours. The 4,6-substituted tryptophan of structure (6) is recoveredfrom the reaction zone by extractive methods as is known in the art.

Starting materials for use in Scheme A are readily available to one ofordinary skill in the art. For example, certain 4,6-substitutedindole-2-carboxylic acids are described in J. Med. Chem. 33 2944-461990.

The following examples present typical syntheses as described in SchemeA. These examples are understood to be illustrative only and are notintended to limit the scope of the present invention in any way. As usedherein, the following terms have the indicated meanings: "g" refers tograms; "mmol" refers to millimoles; "mL" refers to milliliters; "bp"refers to boiling point; "°C." refers to degrees Celsius; "mm Hg" refersto millimeters of mercury; "μL" refers to microliters; "μg" refers tomicrograms; and "μM" refers to micromolar.

EXAMPLE 1 DL-4,6-Dichlorotryptophan

Step a: 4,6-Dichloroindole

Dissolve 4,6-dichloro-indole-2-carboxylic acid (1.0 g, 4.35 mmol) inquinoline (25 mL). Add copper powder (100 mg) and heat to 220° C. for 3hours. Pour the resulting black solution into cold concentratedhydrochloric acid (300 mL) and extract into ethyl ether (500 mL).Filter, wash with 1M hydrochloric acid (2×200 mL), water (100 mL) anddry (MgSO₄). Evaporate the solvent in vacuo to give a brown oil (0.76g). Purify by silica gel chromatography (17% ethyl acetate/hexane) togive the title compound as an amber oil (0.66 g, 81%).

¹ H NMR (CDCl₃ /TMS): 6.56-6.59 ppm (1H, m), 7.10-7.13 ppm (2H, m),7.16-7.18 ppm (1H, m). ¹³ C NMR (CDCl₃), ppm: 101.48, 110.16, 120.24,125.77, 125.94, 126.54, 122.79, 106.54.

Step b: Ethyl 2-(hydroxyimino)-3-(4,6-dichloro-3-indolyl)propanoate

Mix 4,6-dichloroindole (5.90 g, 31.72 mmol), potassium carbonate (1.81g, 47.6 mmol) and anhydrous methylene chloride (200 mL). Stir and add asolution of ethyl 3-bromo-2-hydroxyiminopropanoate (7.00 g, 33.31 mmol)in methylene chloride (75 mL). Stir under a nitrogen atmosphere for 48hours. Take the solution up in methylene chloride (100 mL) and wash withwater (300 mL), saturated sodium hydrogen carbonate (200 mL) and brine(100 mL). Dry (MgSO₄) and evaporate the solvent in vacuo to give a tansolid. Purify by silica gel chromatography (1 to 3% acetone inchloroform) to give the title compound as a yellow solid (7.00 g, 99%based on consumed starting material). Recrystallize (diethylether/hexane) to give the title compound as white crystals (4.0 g, 56%);mp 175°-176° C.

¹ H NMR (DMSO-d₆ /TMS): 1.19 ppm (3H, t), 4.15 ppm (2H, q), 4.15 ppm(2H, s), 6.95 ppm (1H, s), 7.10 ppm (1H, d), 7.40 ppm (1H, d), 11.3 ppm(1H, bs), 12.35 (1H, s).

Anal. Calcd for C₁₃ H₁₂ Cl₂ N₂ O₃ : C 49.54; H, 3.84; N, 8.89; Found: C,49.30; H, 3.78; N, 8.62.

Step c: DL-4,6-Dichlorotryptophan ethyl ester

Dissolve ethyl 2-(hydroxyimino)-3-(4,6-dichloro-3-indolyl)propanoate(1.10 g, 3.65 mmol) in acetic acid (200 mL) and add activated zinc dust(1.25 g, 19.2 mmol). Stir at room temperature for 72 hours. Evaporatethe acetic acid in vacuo to give a white oil. Take the white oil up inethyl acetate (200 mL) and treat with saturated sodium hydrogencarbonate (500 mL). Filter the resulting white precipitate and separatethe organic phase. Wash with saturated sodium hydrogen carbonate (100mL) and brine (100 mL). Dry (MgSO₄) and evaporate the solvent in vacuoto give the title compound as an amber oil (1.02 g, 98%).

¹ H NMR (CDCl₃ /TMS): 1.25 pppm (3H, t), 1.9 ppm (2H, bs), 3.05 ppm (1H,m), 3.55 ppm (1H, m), 3.9 ppm (1H, m), 4.15 ppm (2H, q), 7.05 ppm (1H,s), 7.05 ppm (1H, d), 7.19 ppm (1H, d), 8.70 ppm (1H, bs).

Step d: DL-4,6-Dichlorotryptophan

Mix DL-4,6-dichlorotryptophan ethyl ester (2.37 g, 7.90 mmol) andpyridine (100 mL). Add benzyl chloroformate (2.89 g, 16.98 mmol) andstir for 12 hours. Dilute with ethyl acetate, wash with 1M hydrochloricacid (2×200 mL) and brine (2×200 mL). Dry (MgSO₄) and evaporate thesolvent in vacuo to give a yellow oil (3.24 g). Purify byrecrystallization (hexane/ethyl acetate) to giveDL-4,6-dichlorotryptophan-benzyloxycarbonyl ethyl ester as white needles(2.37 g, 70%); mp 151°-153° C.

¹ H NMR (CDCl₃ /TMS): 1.20 ppm (3H, t), 3.35 ppm (1H, m), 3.60 ppm (1H,dd), 4.15 ppm (2H, q), 4.75 ppm (1H, m), 5.00 ppm (2H, s), 5.35 ppm (1H,d), 7.00 ppm (1H, s), 7.13 ppm (1H, d), 7.3 ppm (6H, m), 8.10 ppm (1H,bs).

Anal. Calcd for C₂₁ H₂₀ Cl₂ N₂ O₄ : C, 57.94; H, 4.63; N, 6.44; Found:C, 57.58; N, 4.33; N,6.35.

Mix DL-4,6-dichlorotryptophan-benzyloxycarbonyl ethyl ester (2.10 g,4.88 mmol) in 1:1 tetrahydrofuran/water (100 mL) and add lithiumhydroxide monohydrate (615 mg, 14.6 mmol). Stir at room temperature for2 hours. Pour into 1M hydrochloric acid (200 mL) and extract with ethylacetate (200 mL). Dry (MgSO₄) and evaporate the solvent in vacuo to giveDL-4,6-dichlorotryptophan-benzyloxycarbonyl as an amber gum (1.90 mg,100%).

¹ H NMR (CDCl₃ /TMS): 3.35 ppm (1H, m), 3.65 ppm (1H, dd), 4.75 ppm (1H,m), 5.05 ppm (2H, s), 5.45 ppm (1H, s), 7.00 ppm (1H, s), 7.13 ppm (1H,d), 7.3 ppm (6H, m), 8.35 ppm (1H, bs).

Dissolve DL-4,6-dichlorotryptophan-benzyloxycarbonyl (1.99 g, 4.88 mmol)in chloroform (100 mL) and add trimethylsilyl iodide (4.88 g, 24.4mmol). Stir at room temperature for 0.5 hours, quench with methanol andevaporate the solvent in vacuo to give a violet oil. Take up the residuein isopropanol (200 mL) containing a small amount of DL-dithiothreitol.Neutralize the resulting pale yellow solution with propylene oxide (1.45g, 25 mmol) to give a white solid (1.15 g, 86%).

FTIR (KBr) cm⁻¹ : 3418 (NH), 3033 (COOH), 1616 (C═O), 1586-1479(Aromatic C═C). ¹ H NMR (CDCl₃ /TFA/TMS): 3.25 ppm (1H, dd), 4.13 ppm(1H, dd), 4.8 ppm (1H, m), 7.15 ppm (1H, d), 7.2 ppm (1H, s), 7.35 ppm(1H, d), 8.5 ppm (1H, bs). ¹³ C NMR (CDCl₃ /TFA) ppm: 27.2, 55.4, 106.5,110.9. 121.7, 122.0, 125.5, 126.9, 129.5, 138.2, 170.25. MS (m/z): 273(M+, 100%), 255, 237, 227.

Anal. Calcd for C₁₁ H₉ Cl₂ N₂ O₂.1/5H₂ O: C, 47.92; H, 3.44; N, 10.16;Found: C, 47.96; H, 3.84; N, 9.97.

EXAMPLE 2 DL-4,6-Dibromotryptophan

Step a: 4,6-Dibromoindole

Dissolve 5,7-dibromo-indole-2-carboxylic acid (1.39 g, 4.35 mmol) inquinoline (25 mL). Add copper powder (100 mg) and heat to 220° C. for 3hours. Pour the resulting black solution into cold concentratedhydrochloric acid (300 mL) and extract into ethyl ether (500 mL).Filter, wash with 1M hydrochloric acid (2×200 mL), water (100 mL) anddry (MgSO₄). Evaporate the solvent in vacuo to give the title compound.

Step b: Ethyl 2-(hydroxyimino)-3-(4,6-dibromo-3-indolyl)propanoate

Mix 4,6-dibromoindole (2.09 g, 7.58 mmol), potassium carbonate (1.57 g,11.4 mmol) and anhydrous methylene chloride (30 mL). Stir and add asolution of ethyl 3-bromo-2-hydroxyiminopropanoate (1.59 g, 7.58 mmol)in methylene chloride (20 mL). Stir under a nitrogen atmosphere for 48hours. Take the solution up in ethyl acetate and wash with water (100mL), saturated sodium hydrogen carbonate (100 mL) and brine (100 mL).Dry (MgSO₄) and evaporate the solvent in in vacuo and purify by silicagel chromatography to give the title compound.

Step c: DL-4,6-Dibromotryptophan ethyl ester

Dissolve ethyl 2-(hydroxyimino)-3-(4,6-dibromo-3-indolyl)propanoate(1.47 g, 3.65 mmol) in acetic acid (200 mL) and add activated zinc dust(1.25 g, 19.2 mmol). Stir at room temperature for 72 hours. Evaporatethe acetic acid in vacuo and take up in ethyl acetate (200 mL). Treatwith saturated sodium hydrogen carbonate (500 mL). Filter the resultingwhite precipitate and separate the organic phase. Wash with saturatedsodium hydrogen carbonate (100 mL) and brine (100 mL). Dry (MgSO₄) andevaporate the solvent in vacuo to give the title compound.

Step d: DL-4,6-Dibromotryptophan

Mix DL-4,6-dibromotryptophan ethyl ester (1.20 g, 3.07 mmol),triethylamine (341 mg, 3.38 mmol) and methylene chloride (50 mL). Addbenzyl chloroformate (692 mg, 4.06 mmol) and stir for 12 hours. Dilutewith methylene chloride, wash with 1M hydrochloric acid (100 mL) andbrine (100 mL). Dry (MgSO₄) and evaporate the solvent in vacuo andpurify by silica gel chromatography to giveDL-4,6-dibromotryptophan-benzyloxycarbonyl ethyl ester.

Mix DL-4,6-dibromotryptophan-benzyloxycarbonyl ethyl ester (440 mg, 0.84mmol) in 1:1 tetrahydrofuran/water (50 mL) and add lithium hydroxidemonohydrate (106 mg, 2.52 mmol). Stir at room temperature for 2 hours.Pour into 1M hydrochloric acid (150 mL) and extract with ethyl acetate(200 mL). Dry (MgSO₄) and evaporate the solvent in vacuo to giveDL-4,6-dibromotryptophan-benzyloxycarbonyl.

Dissolve DL-4,6-dibromotryptophan-benzyloxycarbonyl (407 mg, 0.82 mmol)in chloroform (50 mL) and add trimethylsilyl iodide (656 mg, 3.28 mmol).Stir at room temperature for 1.5 hours and pour the resulting violetsolution into isopropanol (50 mL) containing a small amount ofDL-dithiothreitol. Neutralize with propylene oxide to give a whitesemi-solid. Take this mixture up in 1M hydrochloric acid (200 mL), treatwith charcoal and wash with methylene chloride. Evaporate the water invacuo. Dissolve the solid in methanol (50 mL) and neutralize withpropylene oxide. Add ethyl ether and filter to give the title compound.

The following compounds can be prepared analogously to that described inExample 1-2:

DL-4-Bromo-6-fluorotryptophan;

DL-4-Bromo-6-chlorotryptophan;

DL-4-Ethyl-6-bromotryptophan.

The compounds of Formula Ib may be prepared using techniques andprocedures well known and appreciated by one of ordinary skill in theart. A general synthetic procedure for preparing these compounds is setforth in Scheme B. In Scheme B, all substituents unless otherwiseindicated are as previously defined.

Scheme B provides a general synthetic scheme for preparing compounds ofFormula Ib.

In step a, the appropriate 3,5-disubstituted aniline of structure (7) isiodinated to give the corresponding 2-iodo-3,5-disubstituted-aniline ofstructure (8).

For example, the appropriate 3,5-disubstituted aniline of structure (7)is contacted with a molar equivalent of an appropriate iodinating agentsuch as N-iodosuccinimide. The reactants are typically contacted in asuitable acidic organic solvent such as acetic acid/methylene chloride.The reactants are typically stirred together at room temperature in theabsence of light for a period of time ranging from 5-24 hours. The2-iodo-3,5-disubstituted-aniline of structure (8) is recovered from thereaction zone by extractive methods as is known in the art. It may bepurified by silica gel chromatography.

Alternatively, an appropriate 2-iodo-3,5-disubstituted-aniline ofstructure (8) may be prepared from an appropriate2,4-disubstituted-b-nitroaniline. ##STR5##

First, an appropriate 2,4-disubstituted-6-nitroaniline is iodinated asis known in the art, such as 50% sulfuric acid, sodium nitrite andpotassium iodide, to give the corresponding2-iodo-3,5-disubstituted-nitrobenzene.

Second, the nitro functionality of an appropriate2-iodo-3,5-disubstituted-nitrobenzene is reduced as is known in the art,such as tin (II) chloride dihydrate, to give the corresponding2-iodo-3,5-disubstituted-aniline of structure (8).

In step b, appropriate 2-iodo-3,5-disubstituted-aniline of structure (8)is protected to give the N-protected-3,5-disubstituted-2-iodoaniline ofstructure (9). The selection and utilization of appropriate protectinggroups are well known to one of ordinary skill in the art and aredescribed in "Protective Groups in Organic Synthesis", Theodora W.Greene, Wiley (1981).

In step c, the appropriate N-protected-3,5-disubstituted-2-iodoanilineof structure (9) is converted to the correspondingN-protected-3,5-disubstituted-2-(trimethylstannyl)aniline of structure(10).

For example, the appropriate N-protected-3,5-disubstituted-2-iodoanilineof structure (9) is contacted with a molar excess of an appropriatestannylating agent, such as hexamethylditin, a molar excess of anon-nucleophilic base, such as N-methylmorpholine or sodium hydride, anda catalytic amount of a palladium(0) reagent such astris(dibenzylideneacetone)dipalladium(0) or Pd(CN)₂ Cl₂. The reactantsare typically contaced in a suitable organic solvent such as toluene.The reactants are typically stirred together for a period of timeranging from 10-45 hours and at a temperature range of from 40°-80° C.The N-protected-3,5-disubstituted-2-(trimethylstannyl)aniline ofstructure (10) is recovered from the reaction zone and purified bysilica gel chromatography.

In step d, the appropriateN-protected-3,5-disubstituted-2-(trimethylstannyl)aniline of structure(10) is reacted with (S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazolidineacetylchloride (11) to give the correspondingN-protected-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-disubstituted-anilineof structure (12).

For example, the appropriateN-protected-3,5-disubstituted-2-(trimethylstannyl)aniline of structure(10) is contacted with a molar equivalent of(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazolidineacetyl chloride (11). Thereactants are typically contacted in a suitable organic solvent such astoluene. The reactants are typically stirred together for a period oftime ranging from 1-5 hours and at a temperature range of from roomtemperature to 60° C. TheN-protected-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-disubstituted-anilineof structure (12) is recovered from the reaction zone and purified bysilica gel chromatography.

In step e, the appropriateN-protected-2-[1-oxo-2-[(S)3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-disubstituted-anilineof structure (12) is deprotected to give the4,6-disubstituted-kynurenine of structure (13).

For example, the appropriateN-protected-2-[1-oxo-2[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]3,5-disubstituted-anilineof structure (12) is contacted with a molar excess of trimethylsilyliodide. The reactants are typically contacted in a suitable organicsolvent such as chloroform. The reactants are typically stirred togetherat room temperature for a period of time ranging from 1-5 hours. The4,6-disubstituted-kynurenine of structure (13) is recovered from thereaction zone by extractive methods as is known in the art.

Alternatively, the compounds may be sequentially deprotected with TFA toremove the Boc group, in NaOH to diesterity and TMSI to remove the CBZgroup.

Starting materials for use in Scheme B are readily available to one ofordinary skill in the art. For example,(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazolidineacetyl chloride (11) isdescribed in J. Org. Chem. 53 6138-39 1988.

The following examples present typical syntheses as described in SchemeB. These examples are understood to be illustrative only and are notintended to limit the scope of the present invention in any way.

EXAMPLE 3 L-4,6-Dichlorokynurenine

Step a: 2-Iodo-3,5-dichloroaniline

Dissolve 3,5-dichloroaniline (10 g, 61.5 mmol) in 1:1 aceticacid/methylene chloride (200 mL). Add N-iodosuccinimide (16.9 g, 61.5mmol) as a solid and stir at room temperature in the absence of lightfor 8 hours. Pour the solution into saturated sodium hydrogen carbonate(200 mL) and extract into methylene chloride (100 mL). Wash withsaturated sodium hydrogen carbonate (200 mL), dry (MgSO₄) and evaporatethe solvent in vacuo. Purify by silica gel chromatography to give thetitle compound (8.1 g, 45%); mp 61°-63° C.

¹ H NMR (CDCl₃) 4.4 ppm (2H, bs), 6.70 ppm (1H, d), 6.88 ppm (1H, d); ¹³C NMR (CDCl₃) ppm: 85.77, 111.75, 118.47, 135.21, 139.78, 149.48.

Anal. Calcd for C₆ H₄ Cl₂ IN: C, 25.03; H, 1.40; N, 4.87; Found: C,25.33; H, 1.36; N, 4.87.

Step b: N-(t-butoxycarbonyl)-3,5-dichloro-2-iodoaniline

Mix 2-iodo-3,5-dichloroaniline (11.67 g, 40.5 mmol),di-tert-butyldicarbonate (44 g, 203 mmol) and a catalytic amount ofdimethylaminopyridine. Stir for 4 hours at room temperature, take up inethyl acetate (200 mL), wash with 1M hydrochloric acid (200 mL),saturated sodium chloride (100 mL) and dry (MgSO₄). Evaporate thesolvent in vacuo and purify by silica gel chromatography to giveN-(bis-t-butoxycarbonyl)-3,5-dichloro-2-iodoaniline as a white solid(17.4 g, 87%); mp 131°-132° C.

¹ H NMR (CDCl₃ /TMS) 1.45 ppm (18H, s), 7.11 ppm (1H, d), 7.41 ppm (1H,d); ¹³ C NMR (CDCl₃) ppm: 27.87, 83.22, 103.18, 127.31, 128.07, 134.66,140.08, 144.80, 149.63.

Anal. Calcd for C₁₆ H₂₀ Cl₂ INO₄ : C, 39.37, H, 4.13; N, 2.87; Found: C,39.37; H, 4.20; N, 3.12.

Mix N-(bis-t-butoxycarbonyl)-3,5-dichloro-2-iodoaniline (17.39 g, 35.64mmol), potassium carbonate (6.4 g, 46 mmol) and ethanol (200 mL). Heatfor 5 hours, pour into water (200 mL), extract into ethyl acetate (150mL) and dry (MgSO₄). Evaporate the solvent in vacuo and purify by silicagel chromatography (10:1 hexane/ethyl acetate) to give the titlecompound as a white crystaline solid (12.56 g, 91%); mp 64°-66° C. ¹ HNMR (CDCl₃ /TMS) 1.60 ppm (9H, s), 7.1 ppm (1H, bs), 7.19 ppm (1H, d),8.12 ppm (1H, d).

Step c: N-(t-butoxycarbonyl)-3,5-dichloro-2-(trimethylstannyl)aniline

Mix sodium hydride (413 mg of a 60% suspension in mineral oil, 10.32mmol) and 1-methyl-2-pyrrolidinone (5 mL). Place under an argonatmosphere and cool to 0° C. Add a solution ofN-(t-butoxycarbonyl)-3,5-dichloro-2-iodoaniline (3.33 g, 8.60 mmol) in1-methyl-2-pyrrolidinone (50 mL). Stir at 0° C. for 15 minutes, then for30 minutes at room temperature. Cool to 0° C. and add hexamethylditin(6.7 g, 21 mmol) followed by Pd(CN)₂ Cl₂ (223 mg). Stir for 5 minutes,dilute the resulting black solution with diethyl ether (100 mL) and passthrough a bed of Celite filter aid. Wash the solution with saturatedsodium chloride (2×100 mL), water (100 mL) and dry (MgSO₄). Evaporatethe solvent in vacuo and purify by silica gel chromatography (14:1hexane/ethyl acetate) to give the title compound as a white crystallinesolid (2.20 g, 60%).

¹ H NMR (CDCl₃ /TMS) 0.50 ppm (9H, s), 1.51 ppm (9H, s) , 6.75 ppm (1H,bs), 7.05 ppm (1H, d), 7.70 ppm (1H, d).

Step d:N-(t-butoxycarbonyl)-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-dichloroaniline

Mix (S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazolidineacetyl chloride (1.85g, 6.21 mmol), powdered 4A molecular sieves (4 g) and toluene (50 mL).Stir under an argon atmosphere for 1 hour. Add a solution ofN-(t-butoxycarbonyl)-3,5-dichloro-2-(trimethylstannyl)aniline (2.20 g,5.18 mmol) in toluene (50 mL) followed by Pd(CN)₂ Cl₂ (134 mg). Heat to80° C. for 1.5 hours, cool and purify directly by silica gelchromatography (3:1 hexane/ethyl acetate) to give the title compound asa colorless oil (1.42 g, 53%).

¹ H NMR (CDCl₃ /TMS) 1.50 ppm (9H, s), 3.50 ppm (1H, dd), 3.8-4.1 ppm(1H, m), 4.5 ppm (1H, bs), 5.25 ppm (2H, dd), 5.35 ppm (1H, dd), 5.5-5.7ppm (1H, bs), 7.05 ppm (1H, bs), 7.45 ppm (5H, s), 7.85 ppm (1H, d),8.15 ppm (1H, d).

Step e: L-4,6-Dichlorokynurenine

DissolveN-(t-butoxycarbonyl)-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-dichloroaniline(700 mg, 1.34 mmol) in methylene chloride (50 mL) and treat withtrifluoroacetic acid (20 mL). Stir for 3 hours, pour into saturatedsodium hydrogen carbonate (100 mL) and extract into methylene chloride(100 mL). Wash with saturated sodium hydrogen carbonate (100 mL),saturated sodium chloride (100 mL) and dry (MgSO₄). Evaporate thesolvent in vacuo and purify by silica gel chromatography (3:1hexane/ethyl acetate) to give2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-dichloroanilineas a pale yellow oil (380 mg, 67%).

¹ H NMR (CDCl₃ /TMS) 3.60 ppm (1H, dd), 4.0-4.3 ppm (1H, m), 4.45 ppm(1H, bs), 4.9-5.3 ppm (2H, bs), 5.25 ppm (2H, dd), 5.45 ppm (1H, dd),5.6 ppm (1H, bs), 6.55 ppm (1H, d), 6.70 ppm (1H, d), 7.45 ppm (5H, s).

Dissolve2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3,5-dichloroaniline(380 mg, 0.90 mmol) in methanol (10 mL ) and treat with 1N sodiumhydroxide (0.99 mL, 0.99 mmol ). Stir for 6 hours, pour into 1Mhydrochloric acid (100 mL) and extract with ethyl acetate (100 mL). Dry(MgSO₄) and evaporate the solvent in vacuo. Purify by silica gelchromatography (1 to 5% methanol in chloroform) to giveN-(benzyloxycarbonyl)-L-4,6-dichlorokynurenine as a yellow oil (157 mg,43%).

¹ H NMR (CDCl₃ /TMS) 3.65 ppm (2H, dd), 4.80 ppm (1H, m), 5.15 ppm (2H,dd), 5.9 ppm (1H, d), 6.0-6.2 ppm (2H, bs), 6.50 ppm (1H, d), 6.70 ppm(1H, d), 7.35 ppm (5H, s).

Dissolve N-(benzyloxycarbonyl)-L-4,6-dichlorokynurenine (157 mg, 0.43mmol) in chloroform (30 mL) and add trimethylsilyl iodide (426 mg, 2.13mmol). Stir under an argon atmosphere for 1 hour, quench with methanoland evaporate the solvent in vacuo. Take up the resulting red oil inisopropanol (10 mL) containing a trace amount of DL-dithiothreitol.Treat the resulting pale yellow solution with propylene oxide (122 mg,2.13 mmol) to give the title compound as a yellow solid. Wash withdiethyl ether (500 mL) and dry at 60° C. under 1 mm Hg to give the titlecompound (67 mg, 57%).

¹ H NMR (DMSO-d6/TMS) 2.90 ppm (1H, dd), 3.20 ppm (1H, dd), 4.85 ppm(1H, dd), 6.6 ppm (1H, d), 6.65 ppm (1H, d), 6.7 ppm (2H, s), 7.5-8.0ppm (3H, bs); MS (FAB) m/e 277 (M+H, 100), 260 (5), 188 (15).

EXAMPLE 4 4-fluoro-6-Bromo-kynurenine

Step a: 2-Iodo-3-bromo-5-fluoroaniline

Mix 4-fluoro-2-nitroaniline (15.6 g, 0.1 mol) in water (400 mL) and add48% hydrobromic acid (1 kg). Add bromine (16 g, 0.1 mol) with stirringand stir for 1 hour. Dilute to 2 L and cool to 7° C. Filter, wash withwater and dry to give fluoro-2-bromo-6-nitroaniline.

Mix 4-fluoro-2-bromo-6-nitroaniline (15.5 g, 66.0 mmol) and 50% sulfuricacid (200 mL) and cool to 0°-5° C. Add, by dropwise addition, a solutionof sodium nitrite (6.0 g, 86.4 mmol) in water (50 mL). Stir for 30minutes and add solid potassium iodide (29 g, 173 mmol). Take up theproduct in ethyl acetate (300 mL) wash with saturated sodium hydrogencarbonate (2×200 mL), saturated sodium metabisulfite (2×200 mL) andwater (200 mL). Dry (MgSO₄), evaporate the solvent in vacuo and purifyby silica gel chromatography to give1-iodo-2-nitro-4-fluoro-6-bromobenzene as a yellow crystalline solid(17.81 g, 76%); mp 75°-76° C.

¹ H NMR (CDCl₃ /TMS) 7.35 ppm (1H, dd), 7.65 ppm (1H, dd); ¹³ C NMR(CDCl₃) ppm: 111.23, 111.58, 122.98, 123.31, 133.82, 133.94.

Anal. Calcd for C₆ H₂ BrFINO₂ : C, 20.83; H, 0.58; N, 4.05; Found: C,21.02; H, 0.74; N, 4.24.

Treat 1-iodo-2-nitro-4-fluoro-6-bromobenzene (5.0 g, 14.46 mmol) withtin (II) chloride dihydrate (12 g, 58 mmol) in refluxing ethanol (150mL). Stir at reflux for 24 hours, pour into a mixture of ethyl acetate(200 mL) and saturated sodium hydrogen carbonate (500 mL). Filter andwash the filtrate with saturated sodium hydrogen carbonate (200 mL) andwater (200 mL). Dry (MgSO₄), evaporate the solvent in vacuo and purifyby silica gel chromatography to give the title compound as an off-whitesolid (3.45 g, 76%).

¹ H NMR (CDCl₃ /TMS) 4.5 ppm (2H, bs), 6.45 ppm (1H, dd), 7.85 ppm (1H,dd).

Step b: N-(t-butoxycarbonyl)-3-bromo-5-fluoro-2-iodoaniline

Mix 2-Iodo-3-bromo-5-fluoroaniline (3.00 g, 9.5 mmol),di-tert-butyldicarbonate (4.15 g, 19 mmol) and a catalytic amount ofdimethylaminopyridine. Stir for 4 hours at room temperature, take up inethyl acetate, wash with 1M hydrochloric acid, saturated sodium chlorideand dry (MgSO₄). Evaporate the solvent in vacuo and purify by silica gelchromatography to giveN-(bis-t-butoxycarbonyl)-3-bromo-5-fluoro-2-iodoaniline as a white solid(4.36 g, 90%).

¹ H NMR (CDCl₃ /TMS) 1.45 ppm (18H, s), 6.95 ppm (1H, dd), 7.40 ppm (1H,dd).

Mix N-(bis-t-butoxycarbonyl)-3-bromo-5-fluoro-2-iodoaniline (4.00 g,7.75 mmol), potassium carbonate (10 mmol) and ethanol (50 mL). Heat for5 hours, pour into water (50 mL), extract into ethyl acetate (40 mL) anddry (MgSO₄). Evaporate the solvent in vacuo and purify by silica gelchromatography to give the title compound as a white crystaline solid(2.66 g, 83%).

¹ H NMR (CDCl₃ /TMS) 1.55 ppm (9H, s), 7.15 ppm (1H, dd), 7.20 ppm (1H,bs), 7.95 ppm (1H, dd).

Step c:N-(t-butoxycarbonyl)-3-bromo-5-fluoro-2-(trimethylstannyl)aniline

Mix sodium hydride (317 mg of a 60% suspension in mineral oil, 7.9 mmol)and 1-methyl-2-pyrrolidinone (5 mL). Place under an argon atmosphere andcool to 0° C. Add a solution ofN-(t-butoxycarbonyl)-3-bromo-5-fluoro-2-iodoaniline (2.50 g, 6.10 mmol)in 1-methyl-2-pyrrolidinone (50 mL). Stir at 0° C. for 15 minutes, thenfor 30 minutes at room temperature. Cool to 0° C. and addhexamethylditin (9.8 g, 30 mmol) followed by Pd(CN)₂ Cl₂ (223 mg). Stirfor 5 minutes, dilute the resulting black solution with diethyl ether(100 mL) and pass through a bed of Celite filter aid. Wash the solutionwith saturated sodium chloride (2×100 mL), water (100 mL) and dry(MgSO₄). Evaporate the solvent in vacuo and purify by silica gelchromatography to give the title compound as a white solid (1.50 g,54%).

¹ H NMR (CDCl₃ /TMS) 0.50 ppm (9H, s), 1.50 ppm (9H, s), 6.80 ppm (1H,bs), 7.00 ppm (1H, dd), 7.65 ppm (1H, dd).

Step d:N-(t-butoxycarbonyl)-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl],3-bromo-5-fluoroaniline

Mix (S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazolidineacetyl chloride (946 g,3.18 mmol), powdered 4A molecular sieves (4 g) and toluene (50 mL). Stirunder an argon atmosphere for 1 hour. Add a solution ofN-(t-butoxycarbonyl)-3-bromo-5-fluoro-2-(trimethylstannyl)aniline (1.44g, 3.18 mmol) in toluene (50 mL) followed by Pd(CN)₂ Cl₂ (134 mg). Heatto 80° C. for 1.5 hours, cool and purify directed by silica gelchromatography (3:1 hexane/ethyl acetate) to give the title compound asa white crystalline solid (123 mg, 7%).

¹ H NMR (CDCl₃ /TMS) 1.55 ppm (9H, s), 3.45-4.25 ppm (2H, m), 4.45 ppm(1H, bs), 5.20 ppm (2H, m), 5.4-5.6 ppm (2H, m), 7.00 ppm (1H, m),7.35-7.45 ppm (6H, bs), 7.90 ppm (1H, m).

Step e: L-6-Bromo-4-fluorokynurenine

DissolveN-(t-butoxycarbonyl)-2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3-bromo-5-fluoroaniline(1.34 mmol) in methylene chloride (50 mL) and treat with trifluoroaceticacid (20 mL). Stir for 3 hours, pour into saturated sodium hydrogencarbonate (100 mL) and extract into methylene chloride (100 mL). Washwith saturated sodium hydrogen carbonate (100 mL), saturated sodiumchloride (100 mL) and dry (MgSO₄). Evaporate the solvent in vacuo andpurify by silica gel chromatography (3:1 hexane/ethyl acetate) to give2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3-bromo-5-fluoroaniline.

Dissolve2-[1-oxo-2-[(S)-3-(benzyloxycarbonyl)-5-oxo-4-oxazoldine-2-yl]ethyl]-3-bromo-5-fluoroaniline(0.90 mmol) in methanol (10 mL) and treat with 1N sodium hydroxide (0.99mL, 0.99 mmol). Stir for 6 hours, pour into 1M hydrochloric acid (100mL) and extract with ethyl acetate (100 mL). Dry (MgSO₄) and evaporatethe solvent in vacuo. Purify by silica gel chromatography (1 to 5%methanol in chloroform) to giveN-(benzyloxycarbonyl)-L-6-bromo-4-fluorokynurenine.

Dissolve N-(benzyloxycarbonyl)-L-6-bromo-4-fluorokynurenine (0.43 mmol)in chloroform (30 mL) and add trimethylsilyl iodide (426 mg, 2.13 mmol).Stir under an argon atmosphere for 1 hour, quench with methanol andevaporate the solvent in vacuo. Take up the resulting residue inisopropanol (10 mL) containing a trace amount of DL-dithiothreitol.Treat the solution with propylene oxide (122 mg, 2.13 mmol) to give thetitle compound.

The following compounds can be prepared analogously to those describedin Examples 3 and 4:

6-Bromo-4-chlorokynurenine;

6-Bromo-4-bromokynurenine;

6-Ethyl-4-bromokynurenine.

An alternate synthetic procedure for preparing compounds of Formula Ibis outlined in Scheme C. In Scheme C, all substituents unless otherwiseindicated, are as previously defined.

Scheme C provides an alternate synthetic procedure for preparingcompounds of Formula Ib.

In step a, the appropriate 4,6-substituted tryptophan ethyl ester ofstructure (5) is protected to give the4,6-substituted-tryptophan-benzyloxycarbonyl ethyl ester of structure(14) as described previously in Scheme A, step d.

In step b, the appropriate 4,6-substituted tryptophan-benzylcarbonylethyl ester of structure (14) is oxidatively cleaved to give theN-(benzyloxycarbonyl)-3,5-disubstituted-kynurenine ethyl ester ofstructure (15).

For example, the appropriate 4,6-substitutedtryptophan-benzyloxycarbonyl ethyl ester of structure (14) is contactedwith a molar excess of an oxidating agent such as 4-t-butyliodylbenzene. The reactants are typically contacted in a suitableorganic solvent such as chlorobenzene. The reactants are typicallystirred together for a period of time ranging from 2-24 hours and at atemperature range of from room temperature to reflux. TheN-(benzyloxycarbonyl)-3,5-disubstituted-kynurenine ethyl ester ofstructure (15) is recovered from the reaction zone by evaporate of thesolvent. It can be purified by silica gel chromatography. ##STR6##

Alternatively, the appropriate 4,6-disubstitutedtryptophan-benzylcarbonyl ethyl ester of structure (14) may beoxidatively cleaved with ozone as is known in the art to give anintermediateN-(benzyloxycarbonyl)-3,5-disubstituted-N-formyl-kynurenine. TheN-formyl functionality of the intermediateN-(benzyloxycarbonyl)-3,5-disubstituted-N-formyl-kynurenine is thenremoved by acid hydrolysis to give the correspondingN-(benzyloxycarbonyl)-3,5-disubstituted-kynurenine ethyl ester ofstructure (15).

In step c, the protecting groups of the appropriateN-(benzyloxycarbonyl)-3,5-disubstituted-kynurenine ethyl ester ofstructure (15) are removed to give the corresponding3,5-disubstituted-kynurenine of structure (13) as described previouslyin Scheme A, step d.

Starting materials for use in Scheme C are readily available to one ofordinary skill in the art. For example, 4-t-butyl iodylbenzene isdescribed in J. Chem. Soc., Chem. Commun. 1887-88.

The following examples present typical syntheses as described in SchemeC. These examples are understood to be illustrative only and are notintended to limit the scope of the present invention in any way.

EXAMPLE 5 4,6-Dibromokynurenine

Step a: N-(Benzyloxycarbonyl)-4,6-dibromotryptophan ethyl ester

Mix DL-4,6-dibromotryptophan ethyl ester (1.20 g, 3.07 mmol),triethylamine (341 mg, 3.38 mmol) and methylene chloride (50 mL). Addbenzyl chloroformate (692 mg, 4.06 mmol) and stir for 12 hours. Dilutewith methylene chloride, wash with 1M hydrochloric acid (100 mL) andbrine (100 mL). Dry (MgSO₄) and evaporate the solvent in vacuo andpurify by silica gel chromatography to give the title compound.

Step b: N-(Benzyloxycarbonyl)-4,6-dibromokynurenine ethyl ester

Dissolve N-(benzyloxycarbonyl)-4,6-dibromotryptophan ethyl ester (1.05g, 2 mmol) in chlorobenzene (8 mL) and mix with 4-t-butyl iodylbenzene(876 g, 3 mmol). Reflux for 4 hours and evaporate the solvent in vacuo.Purify by silica gel chromatography to give the title compound.

Step c: 4,6-Dibromokynurenine

Mix N-(benzyloxycarbonyl)-4,6-dibromokynurenine ethyl ester (444 mg,0.84 mmol) in 1:1 tetrahydrofuran/water (50 mL) and add lithiumhydroxide monohydrate (106 mg, 2.52 mmol). Stir at room temperature for2 hours. Pour into 1M hydrochloric acid (150 mL) and extract with ethylacetate (200 mL). Dry (MgSO₄) and evaporate the solvent in vacuo to giveN-(benzyloxycarbonyl)-4,6-dibromokynurenine.

Dissolve N-(benzyloxycarbonyl)-4,6-dibromokynurenine (410 mg, 0.82 mmol)in chloroform (50 mL) and add trimethylsilyl iodide (656 mg, 3.28 mmol).Stir at room temperature for 1.5 hours and pour the resulting violetsolution into isopropanol (50 mL) containing a small amount ofDL-dithiothreitol. Neutralize with propylene oxide to give a whitesemi-solid. Take this mixture up in 1M hydrochloric acid (200 mL), treatwith charcoal and wash with methylene chloride. Evaporate the water invacuo to give a white solid. Dissolve the white solid in methanol (50mL) and neutralize with propylene oxide. Add ethyl ether and filter togive the title compound.

EXAMPLE 6 4,6-Dichlorokynurenine (See Example 3)

Step b: N-(Benzyloxycarbonyl)-4,6-dichlorokynurenine ethyl ester ester

Dissolve DL-4,6-dichlorotryptophan-benzyloxycarbonyl ethyl ester (1.90g, 4.42 mmol) in methanol (200 mL), cool to -78° C. and treat with ozoneuntil a blue color is observed (approximately 3-5 minutes). Purge withnitrogen gas and quench with dimethylsulfide (10 mL). Evaporate thesolvent in vacuo, take up in diethyl ether and wash with water (2×150mL) and brine (200 mL). Dry (MgSO₄) and evaporate the solvent in vacuoto giveN-(formyl)-2-[ethyl-4-oxo-2-(benzyloxycarbonylamino)butyrate-4-yl]-3,5-dichloroanilineas an amber oil (1.98 g, 96%).

¹ H NMR (CDCl₃ /TMS) 1.25 ppm (3H, t), 3.55 ppm (2H, d), 4.25 (2H, q),5.80 ppm (1H, m), 5.15 ppm (2H, s), 5.80 ppm (1H, bs), 7.19 ppm (1H, d),7.35 ppm (5H, s), 8.30 ppm (1H, d), 8.40 ppm (1H, s), 8.75 ppm (1H, bs).

DissolveN-(formyl)-2-[ethyl-4-oxo-2-(benzyloxycarbonylamino)butyrate-4-yl]-3,5-dichloroaniline(1.94 g, 4.15 mmol) in methanol (200 mL) and treat with 4N hydrochloricacid/dioxane (8.3 mmol). Stir for 1 hour, pour into saturated sodiumhydrogen carbonate (200 mL) and extract into ethyl acetate (150 mL). Dry(MgSO₄) and evaporate the solvent in vacuo to give the title compound asa yellow oil (1.82 g, 100%)

¹ H NMR (CDCl₃ /TMS) 1.35 ppm (3H, t), 3.65 ppm (2H, dd), 4.25 ppm (2H,q), 4.70 ppm (1H, m), 5.15 ppm (4H, bs), 5.80 ppm (1H, d), 6.55 ppm (1H,d), 6.70 ppm (1H, d), 7.35 ppm (5H, s).

Step c: 4,6-Dichlorokynurenine

Dissolve N-(benzyloxycarbonyl)-4,6-dichlorokynurenine ethyl ester (1.82g, 4.14 mmol) in a 1:1 mixture of tetrahydrofuran/water (100 mL). Treatwith lithium hydroxide monohydrate (522 mg, 12.4 mmol) and warm at 70°C. for 4 hours. Pour into 1M hydrochloric acid (100 mL) and extract intoethyl acetate (150 mL). Dry (MgSO₄) and evaporate the solvent in vacuoto give N-(benzyloxycarbonyl)-4,6dichlorokynurenine as a yellow foam(1.52 g, 89%).

¹ H NMR (CDCl₃ /TMS) 3.50-3.80 ppm (2H, dd), 4.80 ppm (1H, m), 5.15 ppm(2H, bs), 6.0 ppm (1H, d), 6.55 ppm (1H, d), 6.70 ppm (1H, d), 7.00 ppm(3H, bs), 7.35 ppm (5H, s).

Dissolve N-(benzyloxycarbonyl)-4,6-dichlorokynurenine (1.52 g, 3.70mmol) in chloroform (100 mL) and add trimethylsilyl iodide (3.7 g, 18.5mmol). Stir at room temperature under an argon atmosphere for 1 hour,quench with methanol and evaporate the solvent in vacuo. Take up theresulting red oil into isopropanol (10 mL) containing a small amount ofDL-dithiothreitol. Neutralize the resulting pale yellow solution withpropylene oxide (1.0 g, 18.5 mmol) to give a yellow solid. Wash withdiethyl ether (500 mL) and dry at 60° C. under 1 mm Hg to give the titlecompound (870 mg, 85%).

¹ H NMR (DMSO-d6/TMS) 2.90 ppm (1H, dd), 3.15-3.25 ppm (1H, m), 3.85 ppm(1H, dd), 6.60 ppm (1H, d), 6.70 ppm (1H, d), 6.75 ppm (2H, s), 7.6-7.9ppm (3H, bs); MS (FAB) m/e 277 (M+H, 100), 260 (15), 188 (55); HRMS(FAB) Calcd. for C₁₀ H₁₁ Cl₂ N₂ O₃ : M+H 277.01467. Found: M+H 277.0160.

The compounds of Formula IIa and IIb are known in the art. Methods forpreparing these compounds are known in the art.

As noted above, the compounds of Formula Ia, Ib, IIa, and IIb(hereinafter "the compounds") antagonize the effects which excitatoryamino acids have upon the NMDA receptor complex. This antagonist effectcan be demonstrated by their ability to prevent NMDA-stimulated cyclicGMP accumulation in neonatal rat cerebellar tissue. This test is basedupon the phenomenon that when samples of neonatal rat cerebellar tissueare exposed to the agonist, NMDA, there is an increase in cyclic GMPlevels within this tissue. NMDA antagonists inhibit or decrease thisrise in cyclic GMP levels. This test can be performed by methods similarto those of Baron et al., J. Pharmacol. Exp. Ther. Vol 250 page 162(1989).

The compounds exhibit anti-convulsant properties and are useful in thetreatment of epilepsy. They are useful in the treatment of grand malseizures, petit mal seizures, psychomotor seizures, autonomic seizures,etc. One method of demonstrating their anti-epileptic properties is bytheir ability to inhibit the seizures that are caused by theadministration of quinolinic acid, an NMDA agonist. This test can beconducted in the following manner.

One group containing ten mice are administered 0.01-100 μg of testcompound intracerebroventricularly in a volume of 5 microliters ofsaline. A second control group containing an equal number of mice areadministered an equal volume of saline as a control. Approximately 5minutes later, both groups are administered 7.7 micrograms of quinolinicacid intracerebroventricularly in a volume of 5 microliters of saline.The animals are observed for 15 minutes thereafter for signs ofclonic-tonic seizures. The control group will have a statisticallyhigher rate of clonic-tonic seizures than will the test group.

The compounds are useful for preventing or minimizing the damage whichnervous tissues contained within the CNS suffer upon exposure to eitherischemic, hypoxic, or hypoglycemic conditions or as the result ofphysical trauma. Representative examples of such conditions includestrokes or cerebrovascular accidents, hyperinsulinemia, cardiac arrest,physical trauma, drownings, suffocation, and neonatal anoxic trauma. Thecompounds should be administered to the patient within 24 hours of theonset of the hypoxic, ischemic, or hypoglycemic condition in order forthe compounds to effectively minimize the CNS damage which the patientwill experience.

The compounds are also useful in the treatment of neurodegenerativediseases such as Huntington's disease, Alzheimer's disease, seniledementia, glutaric acidaemia type I, Parkinson's disease, multi-infarctdementia, and neuronal damage associated with uncontrolled seizures. Theadministration of these compounds to a patient experiencing such acondition will serve to either prevent the patient from experiencingfurther neurodegeneration or it will decrease the rate at which theneurodegeneration occurs.

As is apparent to those skilled in the art, the compounds will notcorrect any CNS damage that has already occurred as the result of eitherdisease, or a lack of oxygen or sugar. As used in this application, theterm "treat" refers to the ability of the compounds to prevent furtherdamage or delay the rate at which any further damage occurs.

The compounds exhibit an anxiolytic effect and are thus useful in thetreatment of anxiety. The compounds also exhibit an analgesic effect andare useful in controlling pain. The compounds may be co-administeredwith a narcotic analgesic such as morphine, demerol, etc. In addition tolowering the dose of narcotic required, the compounds decrease the rateat which patients develop tolerance to the pharmacological effects ofthese narcotics. It is also believed that this co-administration willhelp to prevent the patient from becoming addicted to the narcotic. Thecompounds are also effective in the treatment of migraine. They can beused prophylactically or to relieve the symptoms associated with amigraine episode.

In order to exhibit these therapeutic properties, the compounds need tobe administered in a quantity sufficient to inhibit the effect which theexcitatory amino acids have upon the NMDA receptor complex. The dosagerange at which these compounds exhibit this antagonistic effect can varywidely depending upon the particular disease being treated, the severityof the patient's disease, the patient, the particular compound beingadministered, the route of administration, and the presence of otherunderlying disease states within the patient, etc. Typically thecompounds exhibit their therapeutic effect at a dosage range of fromabout 0.1 mg/kg/day to about 100 mg/kg/day for any of the diseases orconditions listed above. Repetitive daily administration may bedesirable and will vary according to the conditions outlined above.

It has been discovered that probenecid will potentiate the therapeuticactivity of the excitatory amino acid antagonists of the presentinvention. Thus the compounds will exhibit their therapeutic effects atlower doses and for longer periods in patients who are concurrentlyreceiving probenecid. The mechanism by which probenecid potentiatestheir effects is not fully understood, however it is believed thatprobenecid decreases the rate at which the compounds are removed fromthe central nervous system as well as decreasing the rate of excretionby the kidneys. Probenecid increases the effective concentration ofthese compounds in both the CNS and in the systemic circulation.

Probenecid is known in the art. It is available commercially from MerckSharp and Dohme under the tradename Benemid® as well as being availablefrom numerous other sources. Probenecid is a uricosuric agent and isutilized in the treatment of gout. Probenecid is a renal tubulartransport blocking agent and has been utilized to increase plasma levelsof penicillin. The pharmacology of probenecid is described in detail inthe 45th Edition of the Physicians Desk reference on page 1379.Probenecid is currently available commercially as tablets. The sodiumsalt of probenecid is readily water soluble and injectable dosage fromcan be prepared from this salt using techniques well known to thoseskilled in the art.

The compounds of the invention may be administered concurrently withprobenecid in order to treat any of the diseases or conditions describedabove. The quantity of probenecid that is required to potentiate thetherapeutic effects of the compounds can vary widely depending upon theparticular compound being administered, the patient, and the presence ofother underlying disease states within the patient, etc. Typicallythough, the probenecid may be administered at a dosage of from 0.5-3g/day. Repetitive daily administration may be desirable and will varyaccording to the conditions outlined above. The probenecid willtypically be administered from 2-4 times daily.

With the concurrent administration of probenecid, the dosage range forthe excitatory amino antagonists may be adjusted lower by a factor offrom 2-10. Alternatively, the compounds of Formulae I or II may beadministered at the same dosage range in order to obtain an enhancedeffect due to the higher therapeutic concentrations obtained.

The compounds of the present invention may be administered by a varietyof routes. They are effective if administered orally. The compounds mayalso be administered parenterally (i.e. subcutaneously, intravenously,intramuscularly, intraperitoneally, or intrathecally).

Pharmaceutical compositions can be manufactured utilizing techniquesknown in the art. Typically an antagonistic amount of the compound willbe admixed with a pharmaceutically acceptable carrier.

For oral administration, the compounds can be formulated into solid orliquid preparations such as capsules, pills, tablets, lozenges, melts,powders, suspensions, or emulsions. Solid unit dosage forms can becapsules of the ordinary gelatin type containing, for example,surfactants, lubricants and inert fillers such as lactose, sucrose, andcornstarch or they can be sustained release preparations.

In another embodiment, the compounds can be tableted with conventionaltablet bases such as lactose, sucrose, and cornstarch in combinationwith binders, such as acacia, cornstarch, or gelatin, disintegratingagents such as potato starch or alginic acid, and a lubricant such asstearic acid or magnesium stearate. Liquid preparations are prepared bydissolving the active ingredient in an aqueous or non-aqueouspharmaceutically acceptable solvent which may also contain suspendingagents, sweetening agents, flavoring agents, and preservative agents asare known in the art.

For parenteral administration the compounds may be dissolved in aphysiologically acceptable pharmaceutical carrier and administered aseither a solution or a suspension. Illustrative of suitablepharmaceutical carriers are water, saline, dextrose solutions, fructosesolutions, ethanol, or oils of animal, vegetative, or synthetic origin.The pharmaceutical carrier may also contain preservatives, buffers,etc., as are known in the art. When the compounds are being administeredintrathecally, they may also be dissolved in cerebrospinal fluid as isknown in the art.

The compounds of Formulae Ia, Ib, IIa, or IIb, and the probenecid can beadministered as two different pharmaceutical dosage forms.Alternatively, in order to increase patient convenience, the compoundsand the probenecid may be compounded into a single pharmaceutical dosageform. These pharmaceutical compositions can be manufactured utilizingtechniques known in the art similar to those described above. Typicallyan antagonistic amount of the compound of Formula I and an effectiveamount of probenecid will be admixed with a pharmaceutically acceptablecarrier.

As used in this application:

a) the term "patient" refers to warm blooded animals such as, forexample, guinea pigs, mice, rats, cats, rabbits, dogs, monkeys,chimpanzees, and humans;

b) the term "treat" refers to the ability of the compounds to eitherrelieve, alleviate, or slow the progression of the patient's disease orprophylactically prevent its occurrence or the manifestation of itssymptoms;

c) the phrase "antagonize the effects of excitatory amino acids" and thephrase "excitatory amino acid antagonist" should be referred to theability of the compounds to inhibit or decrease the rate at whichglutamate or glycine produce neurotransmission at the NMDA receptorcomplex, and;

d) the term "neurodegeneration" refers to a progressive death anddisappearance of a population of nerve cells occurring in a mannercharacteristic of a particular disease state and leading to braindamage;

e) the phrase "concurrent administration" refers to administering theprobenecid at an appropriate time so that it will potentiate theantagonistic effects of the compounds of Formula I, Ib, IIa, or IIb.This may means simultaneous administration or administration atappropriate but different times. Establishing such a proper dosingschedule will be readily apparent to one skilled in the art.

The compounds may also be admixed with any inert carrier and utilized inlaboratory assays in order to determine the concentration of thecompounds within the serum, urine, etc., of the patient as is known inthe art.

Neurodegenerative diseases are typically associated with a dysfunctionof NMDA receptors. Thus, the compounds of Formula I may be utilized indiagnostic procedures to aid physicians with the diagnosis ofneurodegenerative diseases. The compounds may be labeled with imagingagents known in the art such as isotopic atoms and administered to apatient in order to determine whether the patient is exhibiting adecreased number of NMDA receptors and the rate at which that loss isoccurring.

What is claimed is:
 1. A method for antagonizing the effects ofexcitatory amino acids upon the NMDA receptor complex comprisingadministering to a patent in need thereof, an antagonistic amount of acompound of the formula: ##STR7## or a pharmaceutically acceptable saltthereof; in which Hal is a halogen atom.
 2. A method for the treatmentof epilepsy comprising administering to a patient in need thereof ananti-epileptic amount of a compound according to claim
 1. 3. A methodfor the treatment of neurodegenerative diseases comprising administeringto a patient in need thereof an effective amount of a compound accordingto claim
 1. 4. A method for preventing ischemic/hypoxic/hypoglycemicdamage to cerebral tissue comprising administering to a patient in needthereof an effective amount of a compound according to claim
 1. 5. Amethod for the treatment of anxiety comprising administering ananxiolytic amount of a compound according to claim
 1. 6. A method forproducing an analgesic effect comprising administering to a patient inneed thereof an analgesic amount of a compound according to claim
 1. 7.A method for the treatment of migraine comprising administering aneffective amount of a compound according to claim
 1. 8. A pharmaceuticalcomposition comprising a compound according to claim 1 in admixture witha pharmaceutically acceptable carrier.
 9. A method for antagonizing theeffects of excitatory amino acids upon the NMDA receptor complexcomprising contacting the NMDA receptor of a patient in need thereofwith an antagonistic amount of a compound according to claim
 1. 10. Amethod according to claim 1 in which Hal is chlorine.